Here we wish to focus attention on the role of DNA supercoiling in chromatin structure and gene expression. We have selected the powerful biological system of Saccharomyces cerevisiae to address the four following specific aims: (1) To elucidate the effect of DNA template linearization in vivo on transcription. -role of the position of the linearization relative to the polarity of a transcription unit -role of the position of the linearization relative to the positions of potential topological boundaries -minichromosomal versus chromosomal genes -role of telomere seed sequences flanking the site(s) of linearization (2) To delineate the mechanism(s) of nucleosome splitting -role of positive DNA supercoiling -role of torsional stress -role of RNA polymerase traversal -role of a helix tracking protein (3) To ascertain whether a template containing split nucleosomes is more efficiently transcribed compared to one containing whole nucleosomes (4) To determine whether DNA replication is required for the repair of split nucleosomes